O-O Bond Activation (and Formation) at Bimetallic Enzyme Active Sites

The overall goal of this proposal is to understand how dioxygen is activated by nonheme diiron enzymes in metabolically critical transformations. These enzymes perform a remarkable range of functions, including the biosynthesis of DNA (ribonucleotide reductase (RNR)), the hydroxylation of organic substrates (soluble methane monooxygenase (sMMOH), toluene monooxygenase), the hydroxylation of the eukaryotic initiation factor 5a to regulate eukaryotic cell proliferation (human deoxyhypusine hydroxylase (hDOHH)), the biosynthesis of antibiotics (CmlA, CmlI), and the production of biodiesel (cyanobacterial aldehyde deformylating oxygenase (cADO)). Important project goals are to understand the roles of the initial diiron(II,III)- superoxo species and the subsequently formed diiron(III)-peroxo intermediates in substrate oxidation, how the latter can be converted to corresponding high-valent iron-oxo species that often serve as the key oxidants for substrate transformation, and to describe the structural, electronic, and reactivity properties of the high-valent intermediates. These goals will be accomplished by a combination of biochemical and biomimetic approaches. Our biochemical effort focuses on the diferric-peroxo intermediates (P) of hDOHH and CmlI we found to have different core structures from the carboxylate-bridged intermediates of sMMOH and RNR. With as many as four different P species to compare kinetically and spectroscopically, we aim to shed light on how these structural differences lead to the reactions their respective enzymes catalyze. Our biomimetic effort will focus on characterizing the various diiron-O2 intermediates listed above to gain detailed insight into their electronic structures and their oxidative reactivity. For example, can a diiron(II,III)-superoxo species hydroxylate toluene like toluene-4-monooxygenase? Is it possible for a diiron(III)- peroxo species oxidize C–H bonds to model a hydroxylase and also mimic the action of cADO in the oxidative deformylation of aldehydes? What factors favor one reaction over the other? Most importantly, how are diferric- peroxo intermediates converted into the high-valent diiron oxidants that carry out the most difficult substrate oxidations. These latter complexes are also critical to our efforts to clarify the nature of the high-valent diiron core in the methane-hydroxylating enzyme intermediate sMMOH-Q using new spectroscopic techniques. Our biomimetic efforts will be extended to the synthesis of Fe–O–Mn and Fe–O–Ce complexes. The Fe–O–Mn complexes mimic high-valent intermediates of the ribonucleotide reductase from the parasite Chlamydia trachomatis and the related R2lox enzymes found in pathogenic bacteria. Understanding the difference in the reactivity properties of high-valent FeFe and FeMn complexes may contribute to the development of better methods for treating infections from such human pathogens. The Fe–O–Ce complexes will help us understand the O–O bond formation mechanism of iron-catalyzed water oxidation by CeIV, which we propose to be just the reverse of the reaction sequence used by the diiron enzymes for dioxygen activation.